Chemistry Seminar with Dr. Catherine Eichhorn from the University of Nebraska-Lincoln at 4:00pm

Structural plasticity is integral to RNA function; however, there are currently few methods to quantitatively resolve RNAs that have multiple structural states. Here, we integrate solution NMR spectroscopy and chemical probing approaches to visualize a two-state conformational ensemble for the central stem-loop 3 (SL3) of 7SK RNA, a critical element for 7SK RNA function in transcription regulation. We find that the SL3 distal end undergoes slow exchange between two equally populated yet structurally distinct states in both isolated SL3 constructs and full-length 7SK RNA. We rationally designed constructs that lock SL3 into a single state and demonstrate that both NMR and chemical probing data fit to a linear combination of the two states. A survey of vertebrate 7SK RNA sequences show exquisite sequence dependence on the two-state equilibrium. Moreover, we find that changing environmental variables such as temperature and ionic strength shifts the equilibrium between the two states. These results provide new insights into 7SK RNA structural dynamics and demonstrate the utility of integrating chemical probing with NMR spectroscopy to gain quantitative insights into RNA conformational ensembles.